The expression of proteins from various organisms can be accomplished in heterologous host cells to produce active and useful products. Numerous host organisms have been developed for such expression, and each organism has its own particular requirements for successful expression of heterologous proteins. Likewise, numerous different expression vectors or constructs have been devised to allow for expression of proteins of interest in host organisms of interest.
Expression of a heterologous protein in a host cell may require that the nucleotide sequence encoding the protein be: 1) suitable for expression in the host cell (e.g., have appropriate codon usage rules applied); 2) operably linked to expression control elements that are functional in the host cell (e.g., fused to a promoter that functions in the host cell); and 3) encode a protein that is not toxic to the cell when expressed.
To express a protein that is toxic to a host cell, many solutions have been devised. One solution is to express the protein under tightly controlled conditions such that expression of the protein is totally or nearly totally repressed until an inducer of expression is provided. Some expression systems can be triggered to express large amounts of protein upon introduction of an appropriately controlled inducer. Production of such large amounts of the toxic protein can results in death of the host cell, but not until after adequate amounts of the protein of interest are produced.
Specific expression vectors for expressing toxic proteins recombinantly in host cells have been developed, including incorporation of regulatory sequences in controlling and enhancing expression of recombinant proteins.
There is a continuing need for improved sequences to enable controlled expression of recombinant or toxic proteins. In particular, elements for potent over-expression of mRNA and protein are needed to allow for higher levels of recombinant protein expression and for expression of toxic or recalcitrant proteins in host cells.